Design of Allele-Specific Protein Methyltransferase Inhibitors

• Published in: Journal of the American Chemical Society Vol. 123; no. 47; pp. 11608 - 11613
• Main Authors: Lin, Qing, Jiang, Fanyi, Schultz, Peter G, Gray, Nathanael S
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 28-11-2001
• Subjects: Alleles; Amino Acid Sequence; Binding Sites; Drug Design; Enzyme Inhibitors - chemical synthesis; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - pharmacology; Kinetics; Mutagenesis, Site-Directed; Protein Conformation; Protein-Arginine N-Methyltransferases - antagonists & inhibitors; Protein-Arginine N-Methyltransferases - chemistry; Protein-Arginine N-Methyltransferases - genetics; Protein-Arginine N-Methyltransferases - metabolism; S-Adenosylmethionine - analogs & derivatives; S-Adenosylmethionine - metabolism; Saccharomyces cerevisiae - drug effects; Saccharomyces cerevisiae - enzymology; Sequence Homology, Amino Acid; Substrate Specificity
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja011423j

Protein arginine methyltransferases, which catalyze the transfer of methyl groups from S-adenosylmethionine (SAM) to arginine side chains in target proteins, regulate transcription, RNA processing, and receptor-mediated signaling. To specifically address the functional role of the individual members of this family, we took a “bump-and-hole” approach and designed a series of N 6-substituted S-adenosylhomocysteine (SAH) analogues that are targeted toward a yeast protein methyltransferase RMT1. A point mutation was identified (E117G) in Rmt1 that renders the enzyme susceptible to selective inhibition by the SAH analogues. A mass spectrometry based enzymatic assay revealed that two compounds, N 6-benzyl- and N 6-naphthylmethyl-SAH, can inhibit the mutant enzyme over the wild-type with the selectivity greater than 20. When the E117G mutation was introduced into the Saccharomyces cerevisiae chromosome, the methylation of Npl3p, a known in vivo Rmt1 substrate, could be moderately reduced by N 6-naphthylmethyl-SAH in the resulting allele. In addition, an N 6-benzyl-SAM analogue was found to serve as an orthogonal SAM cofactor. This analogue is preferentially utilized by the mutant methyltransferase relative to the wild-type enzyme with a selectivity greater than 67. This specific enzyme/inhibitor and enzyme/substrate design should be applicable to other members of this protein family and facilitate the characterization of protein methyltransferase function in vivo when combined with RNA expression analysis.